Olfaction is one of the most complex and poorly understood of human sensory systems. From olfactory receptor (OR) activation to perception, there are many steps that still require further investigation. If we can understand how the OR code for individual odorants and mixtures translates into perception then we can exploit this knowledge to bring significant benefit in several areas. These areas include odor modulators like malodor counteractants that block the perception of unpleasant odors, new flavor and fragrance ingredients that replace non-biodegradable or toxic compounds, and odor enhancers that would limit our reliance on difficult to source compounds from natural sources. The ‘olfactory code’ combinatorial paradigm is centered on the observation that any single OR may be activated by multiple odorants, and conversely most odorants are capable of activating several ORs. In the mouse genome there are ˜1,200 distinct ORs. Humans, by contrast, have ˜400. In both cases, the repertoire of ORs is activated by many thousands of odorants in the world, and it is this combinatorial complexity that allows for the breadth of olfactory sensations we can perceive. However, odorants or ligands for only 95 mouse (˜8%) and 41 human ORs (˜10%) have been identified as of 2014 using traditional deorphanization methods. In addition, the physiological relevance of most ligands for the human ORs, essentially identified in vitro, has not been tested.
Different OR de-orphanization methods have been described in the literature [e.g. Touhara (2007) Deorphanizing vertebrate olfactory receptors: Recent advances in odorant-response assays Neurochem Int 51, 132-139, Saito et al (2009) Odor coding by a Mammalian receptor repertoire. Sci Signal 2, ra9, and Peterlin et al. (2014), The State of the Art of Odorant Receptor Deorphanization: a Report from the Orphanage, J Gen Physiol; 143(5): 527-42]. Many of these methods rely exclusively on cell-based assays where the OR is expressed in non-olfactory cells that are suitable for high-throughput screening. However, ORs are often retained in the endoplasmic reticulum of such heterologous cells. Failing to traffic to the cell surface, the ORs are thus unable to interact with the odorant [Min et al. (2004) Endoplasmic reticulum degradation impedes olfactory G-protein coupled receptor functional expression. BMC Cell Biol 5, 34]. Thus, a systematic approach where hundreds to thousands of different cell lines, where each cell line possess a unique OR protein that can be assessed for odorant activity, is not a suitable approach for comprehensive decoding of the combinatoral interactions between odorants and ORs since many or most of the receptors do not function properly in such cell lines. There is therefore a need for new methods that can rapidly and reliably identify the relatively small subset of ORs, within the entire repertoire of ORs that exist in an organism, that are specifically activated or inhibited by one or more odorants. There is a further need for a method to involve the identification of ORs from the olfactory neurons themselves, where the ORs are presumed to be fully functional, thus circumventing the well-known challenges of OR assays in non-olfactory cells.
Malodor compounds such as indole, skatole (3-methyl indole), and p-cresol generate unpleasant odors that arise for example from latrines and other “bathroom” sources that contain fecal matter. Hence, malodor counteractants that mask or reduce the perceived intensity or modify the perceived quality for example of human smell of the compounds are desirable. Odorant receptors and more particularly malodor receptors have a need to be identified. Receptors that bind to indole and skatole have also been identified and compounds that bind to those receptors have been discovered and reported as potential modulators of malodor. However, the rapid identification of the complete repertoire of receptors that bind to malodors, particularly indole or skatole receptors continues to be desirable due to the numerous ORs that exist in mammals. Assays that rely on new malodor receptors to identify new potent compounds that bind to these receptors are further desired.